PROJECT SUMMARY/ABSTRACT Inflammatory bowel diseases (IBD) result from the convergence of host, environmental, and microbial factors, each necessary, but not sufficient to cause disease. Yet, significant gaps in knowledge remain ? among them, what underlies disease susceptibility and precipitates the onset of IBD. The need to better understand the causes that lead to these diseases remains a challenge. Post disease onset, numerous confounders and complications make it problematic to unravel the myriad of contributing factors. Not surprisingly, most human subject studies fail to go beyond description and association. Inability to sort out cause-effect relationship on a temporal basis further hinders efforts to improve clinical management and outcomes and to develop effective strategies for risk stratification and disease prevention. To address this issue, we propose a multi-disciplinary team approach that will use the lens of a unique clinical model to gain transformative insights that will help move the needle in IBD. The model involves a subset of patients with medically-refractory ulcerative colitis (UC) who have undergone total proctocolectomy with ileal pouch anal anastomosis (UC-IPAA) and who can be followed before and after disease development and serve as their own controls. Half of these patients will develop an inflammatory condition of their ileal pouch within two years (pouchitis). In an initial exploratory study of these patients, two discoveries were made which led to the following hypotheses: [1] IBD patients exhibit an anomalous transcriptional response to microbiota-derived signals that renders them susceptible to the development of pouchitis, but is not sufficient to cause disease. [2] Specific mucosal pathobionts that emerge through selection or acquisition of virulence factors trigger pouchitis on the patient's background of genetic susceptibility. With regard to the latter, differences in specific Bacteroides fragilis capsular polysaccharide (CPS) biosynthetic gene clusters among luminal and mucosal strains may transform a commensal microbe to a pathobiont. Three aims are proposed: (1) to investigate the stimuli, role, and functional impact of specific host genes of the anomalous UC pouch transcriptome that may promote disease susceptibility, by employing single-cell approaches to define cell-type and -specific contributions and mechanisms; (2) to identify and isolate microbes involved in promoting pouchitis including CPS variants and determine the their functional impact on host epithelia and immune cells and role and contribution to initiation of disease, (3) to develop a research resource for the broader community to advance discovery-based or hypothesis-generating science. The study will be conducted by a multi-disciplinary team of experts with a proven and successful record of collaboration and synergy. The proposal links basic, translational, and clinical scientific research by focusing on molecular and genetic mechanisms of host and microbial cells with the objective of building a base for future interventions for many types of complex immune diseases; ultimately, we will be able to alter the outcome for these patients.